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Medicinal Research Reviews Mar 2016Bax, a central cell death regulator, is an indispensable gateway to mitochondrial dysfunction and a major proapoptotic member of the B-cell lymphoma 2 (Bcl-2) family... (Review)
Review
Bax, a central cell death regulator, is an indispensable gateway to mitochondrial dysfunction and a major proapoptotic member of the B-cell lymphoma 2 (Bcl-2) family proteins that control apoptosis in normal and cancer cells. Dysfunction of apoptosis renders the cancer cell resistant to treatment as well as promotes tumorigenesis. Bax activation induces mitochondrial membrane permeabilization, thereby leading to the release of apoptotic factor cytochrome c and consequently cancer cell death. A number of drugs in clinical use are known to indirectly activate Bax. Intriguingly, recent efforts demonstrate that Bax can serve as a promising direct target for small-molecule drug discovery. Several direct Bax activators have been identified to hold promise for cancer therapy with the advantages of specificity and the potential of overcoming chemo- and radioresistance. Further investigation of this new class of drug candidates will be needed to advance them into the clinic as a novel means to treat cancer.
Topics: Amino Acid Sequence; Animals; Humans; Models, Biological; Molecular Sequence Data; Neoplasms; Signal Transduction; Small Molecule Libraries; bcl-2-Associated X Protein
PubMed: 26395559
DOI: 10.1002/med.21379 -
Proceedings of the National Academy of... Sep 2021Regulation of apoptosis is tightly linked with the targeting of numerous Bcl-2 proteins to the mitochondrial outer membrane (MOM), where their activation or inhibition...
Regulation of apoptosis is tightly linked with the targeting of numerous Bcl-2 proteins to the mitochondrial outer membrane (MOM), where their activation or inhibition dictates cell death or survival. According to the traditional view of apoptotic regulation, BH3-effector proteins are indispensable for the cytosol-to-MOM targeting and activation of proapoptotic and antiapoptotic members of the Bcl-2 protein family. This view is challenged by recent studies showing that these processes can occur in cells lacking BH3 effectors by as yet to be determined mechanism(s). Here, we exploit a model membrane system that recapitulates key features of MOM to demonstrate that the proapoptotic Bcl-2 protein BAX and antiapoptotic Bcl-xL have an inherent ability to interact with membranes in the absence of BH3 effectors, but only in the presence of cellular concentrations of Mg/Ca Under these conditions, BAX and Bcl-xL are selectively targeted to membranes, refolded, and activated in the presence of anionic lipids especially the mitochondrial-specific lipid cardiolipin. These results provide a mechanistic explanation for the mitochondrial targeting and activation of Bcl-2 proteins in cells lacking BH3 effectors. At cytosolic Mg levels, the BH3-independent activation of BAX could provide localized amplification of apoptotic signaling at regions enriched in cardiolipin (e.g., contact sites between MOM and mitochondrial inner membrane). Increases in MOM cardiolipin, as well as cytosolic [Ca] during apoptosis could further contribute to its MOM targeting and activity. Meanwhile, the BH3-independent targeting and activation of Bcl-xL to the MOM is expected to counter the action of proapoptotic BAX, thereby preventing premature commitment to apoptosis.
Topics: Cardiolipins; Cell Membrane Permeability; Cytosol; Humans; Mitochondrial Membrane Transport Proteins; Mitochondrial Membranes; bcl-2-Associated X Protein; bcl-X Protein
PubMed: 34493661
DOI: 10.1073/pnas.2025834118 -
Scientific Reports Feb 2021The initiation of apoptosis is a core mechanism in cellular biology by which organisms control the removal of damaged or unnecessary cells. The irreversible activation...
The initiation of apoptosis is a core mechanism in cellular biology by which organisms control the removal of damaged or unnecessary cells. The irreversible activation of caspases is essential for apoptosis, and mathematical models have demonstrated that the process is tightly regulated by positive feedback and a bistable switch. BAX and SMAC are often dysregulated in diseases such as cancer or neurodegeneration and are two key regulators that interact with the caspase system generating the apoptotic switch. Here we present a mathematical model of how BAX and SMAC control the apoptotic switch. Formulated as a system of ordinary differential equations, the model summarises experimental and computational evidence from the literature and incorporates the biochemical mechanisms of how BAX and SMAC interact with the components of the caspase system. Using simulations and bifurcation analysis, we find that both BAX and SMAC regulate the time-delay and activation threshold of the apoptotic switch. Interestingly, the model predicted that BAX (not SMAC) controls the amplitude of the apoptotic switch. Cell culture experiments using siRNA mediated BAX and SMAC knockdowns validated this model prediction. We further validated the model using data of the NCI-60 cell line panel using BAX protein expression as a cell-line specific parameter and show that model simulations correlated with the cellular response to DNA damaging drugs and established a defined threshold for caspase activation that could distinguish between sensitive and resistant melanoma cells. In summary, we present an experimentally validated dynamic model that summarises our current knowledge of how BAX and SMAC regulate the bistable properties of irreversible caspase activation during apoptosis.
Topics: Antineoplastic Agents; Apoptosis; Apoptosis Regulatory Proteins; Caspases; HeLa Cells; Humans; Melanoma; Mitochondrial Proteins; Models, Biological; bcl-2-Associated X Protein
PubMed: 33558564
DOI: 10.1038/s41598-021-82215-2 -
Biochimica Et Biophysica Acta Mar 2016Pore-forming proteins and peptides act on their targeted lipid bilayer membranes to increase permeability. This approach to the modulation of biological function is... (Review)
Review
Pore-forming proteins and peptides act on their targeted lipid bilayer membranes to increase permeability. This approach to the modulation of biological function is relevant to a great number of living processes, including; infection, parasitism, immunity, apoptosis, development and neurodegeneration. While some pore-forming proteins/peptides assemble into rings of subunits to generate discrete, well-defined pore-forming structures, an increasing number is recognised to form pores via mechanisms which co-opt membrane lipids themselves. Among these, membrane attack complex-perforin/cholesterol-dependent cytolysin (MACPF/CDC) family proteins, Bax/colicin family proteins and actinoporins are especially prominent and among the mechanisms believed to apply are the formation of non-lamellar (semi-toroidal or toroidal) lipidic structures. In this review I focus on the ways in which lipids contribute to pore formation and contrast this with the ways in which lipids are co-opted also in membrane fusion and fission events. A variety of mechanisms for pore formation that involve lipids exists, but they consistently result in stable hybrid proteolipidic structures. These structures are stabilised by mechanisms in which pore-forming proteins modify the innate capacity of lipid membranes to respond to their environment, changing shape and/or phase and binding individual lipid molecules directly. In contrast, and despite the diversity in fusion protein types, mechanisms for membrane fusion are rather similar to each other, mapping out a pathway from pairs of separated compartments to fully confluent fused membranes. Fusion proteins generate metastable structures along the way which, like long-lived proteolipidic pore-forming complexes, rely on the basic physical properties of lipid bilayers. Membrane fission involves similar intermediates, in the reverse order. I conclude by considering the possibility that at least some pore-forming and fusion proteins are evolutionarily related homologues. This article is part of a Special Issue entitled: Pore-Forming Toxins edited by Mauro Dalla Serra and Franco Gambale.
Topics: Animals; Cell Membrane; Colicins; Humans; Membrane Fusion; Membrane Lipids; Perforin; bcl-2-Associated X Protein
PubMed: 26654785
DOI: 10.1016/j.bbamem.2015.11.026 -
International Journal of Molecular... Oct 2022The Bax protein is a pro-apoptotic protein belonging to the Bcl-2 family, involved in inducing apoptosis at the mitochondrial level. Regulating the protein levels of Bax...
The Bax protein is a pro-apoptotic protein belonging to the Bcl-2 family, involved in inducing apoptosis at the mitochondrial level. Regulating the protein levels of Bax is essential to enhancing apoptosis. In the current study, we ascertained the presence of deubiquitinating enzymes (DUBs) associated with Bax by performing the yeast two-hybrid screening (Y2H). We determined that ubiquitin-specific protease 12 (USP12), one of the DUBs, is associated with Bax. The binding of USP12 to Bax shows the interaction as a DUB, which regulates ubiquitination on Bax. Taken together, we believe that USP12 regulates Bax by detaching ubiquitin on K63-linked chains, indicating that USP12 affects the cellular functions of Bax, but it is not related with proteasomal degradation. The half-life of the Bax protein was determined by performing the site-directed mutagenesis of putative ubiquitination sites on Bax (K128R, K189R, and K190R). Of these, Bax (K128R and K190R) showed less ubiquitination; therefore, we compared the half-life of Bax (WT) and Bax K mutant forms in vitro. Interestingly, Bax (K189R) showed a higher ubiquitination level and shorter half-life than Bax (WT), and the (K128R and K190R) mutant form has a longer half-life than Bax (WT).
Topics: bcl-2-Associated X Protein; Apoptosis Regulatory Proteins; Ubiquitination; Ubiquitin Thiolesterase; Apoptosis
PubMed: 36361894
DOI: 10.3390/ijms232113107 -
Experimental Biology and Medicine... May 2019Bax induces mitochondria-dependent programed cell death. While cytotoxic drugs activating Bax have been developed for cancer treatment, clinically effective therapeutics... (Review)
Review
Bax induces mitochondria-dependent programed cell death. While cytotoxic drugs activating Bax have been developed for cancer treatment, clinically effective therapeutics suppressing Bax-induced cell death rescuing essential cells have not been developed. This mini-review will summarize previously reported Bax inhibitors including peptides, small compounds, and antibodies. We will discuss potential applications and the future direction of these Bax inhibitors.
Topics: Amino Acid Sequence; Animals; Apoptosis; Brain Ischemia; Cell-Penetrating Peptides; Cells, Cultured; Disease Models, Animal; Drug Carriers; Drug Design; Humans; Immunoglobulin Fab Fragments; Ku Autoantigen; Mice; Mice, Knockout; Mitochondria; Mitochondrial Membrane Transport Proteins; Mitochondrial Membranes; Mitochondrial Permeability Transition Pore; Neurodegenerative Diseases; Organ Preservation; Pinocytosis; Protein Multimerization; Pulmonary Fibrosis; Rats; Retinal Degeneration; bcl-2-Associated X Protein
PubMed: 30836793
DOI: 10.1177/1535370219833624 -
Cancer Discovery Mar 2022Cancer cell metabolism is increasingly recognized as providing an exciting therapeutic opportunity. However, a drug that directly couples targeting of a metabolic...
UNLABELLED
Cancer cell metabolism is increasingly recognized as providing an exciting therapeutic opportunity. However, a drug that directly couples targeting of a metabolic dependency with the induction of cell death in cancer cells has largely remained elusive. Here we report that the drug-like small-molecule ironomycin reduces the mitochondrial iron load, resulting in the potent disruption of mitochondrial metabolism. Ironomycin promotes the recruitment and activation of BAX/BAK, but the resulting mitochondrial outer membrane permeabilization (MOMP) does not lead to potent activation of the apoptotic caspases, nor is the ensuing cell death prevented by inhibiting the previously established pathways of programmed cell death. Consistent with the fact that ironomycin and BH3 mimetics induce MOMP through independent nonredundant pathways, we find that ironomycin exhibits marked in vitro and in vivo synergy with venetoclax and overcomes venetoclax resistance in primary patient samples.
SIGNIFICANCE
Ironomycin couples targeting of cellular metabolism with cell death by reducing mitochondrial iron, resulting in the alteration of mitochondrial metabolism and the activation of BAX/BAK. Ironomycin induces MOMP through a different mechanism to BH3 mimetics, and consequently combination therapy has marked synergy in cancers such as acute myeloid leukemia. This article is highlighted in the In This Issue feature, p. 587.
Topics: Apoptosis; Cell Death; Humans; Iron; Mitochondria; bcl-2 Homologous Antagonist-Killer Protein; bcl-2-Associated X Protein
PubMed: 34862195
DOI: 10.1158/2159-8290.CD-21-0522 -
Blood Cancer Journal Apr 2023TP53-mutant acute myeloid leukemia (AML) respond poorly to currently available treatments, including venetoclax-based drug combinations and pose a major therapeutic...
TP53-mutant acute myeloid leukemia (AML) respond poorly to currently available treatments, including venetoclax-based drug combinations and pose a major therapeutic challenge. Analyses of RNA sequencing and reverse phase protein array datasets revealed significantly lower BAX RNA and protein levels in TP53-mutant compared to TP53-wild-type (WT) AML, a finding confirmed in isogenic CRISPR-generated TP53-knockout and -mutant AML. The response to either BCL-2 (venetoclax) or MCL-1 (AMG176) inhibition was BAX-dependent and much reduced in TP53-mutant compared to TP53-WT cells, while the combination of two BH3 mimetics effectively activated BAX, circumventing survival mechanisms in cells treated with either BH3 mimetic, and synergistically induced cell death in TP53-mutant AML and stem/progenitor cells. The BH3 mimetic-driven stress response and cell death patterns after dual inhibition were largely independent of TP53 status and affected by apoptosis induction. Co-targeting, but not individual targeting of BCL-2 and MCL-1 in mice xenografted with TP53-WT and TP53-R248W Molm13 cells suppressed both TP53-WT and TP53-mutant cell growth and significantly prolonged survival. Our results demonstrate that co-targeting BCL-2 and MCL-1 overcomes BAX deficiency-mediated resistance to individual BH3 mimetics in TP53-mutant cells, thus shifting cell fate from survival to death in TP53-deficient and -mutant AML. This concept warrants clinical evaluation.
Topics: Animals; Mice; Myeloid Cell Leukemia Sequence 1 Protein; bcl-2-Associated X Protein; Apoptosis; Cell Line, Tumor; Proto-Oncogene Proteins c-bcl-2; Leukemia, Myeloid, Acute; Antineoplastic Agents
PubMed: 37088806
DOI: 10.1038/s41408-023-00830-w -
Cytometry. Part a : the Journal of the... Jan 2010Most cell death in vertebrates proceeds through the intrinsic pathway of apoptosis and results from unregulated increase of mitochondrial membrane permeability.... (Review)
Review
Most cell death in vertebrates proceeds through the intrinsic pathway of apoptosis and results from unregulated increase of mitochondrial membrane permeability. Bcl2-associated X protein (Bax) and Bcl2-antagonist/killer protein (Bak), the effector proapoptotic members of the Bcl-2 family, are, in their active state, the principal accomplices for this permeabilization process. How exactly Bax and Bak are activated has been a matter of major investigation in the last decade, and suitable tools offered by quantitative cytometric methodologies have significantly contributed to the understanding of the function of Bcl-2 family members. Here, we review the most relevant findings in this field and highlight one common trait that has emerged from the diverse new theories: a crucial role in the control of Bax/Bak activation has to be attributed to the BH3-only subset of the Bcl-2 family. BH3-only proteins exert their proapoptotic activity by hierarchical and tightly tuned interactions with other Bcl-2 family members and operate as sensors of intracellular/extracellular death signals and vectors of information to the core apoptotic machinery. Given their essential role in apoptosis, BH3-only molecules are proposed as molecular targets for the cure of diseases associated with abnormal cell death, as in the case with neurodegenerative conditions. As well, they are explored as possible tools for cancer therapy, according to the concept that molecules mimicking the BH3 domain of these proteins could selectively and efficiently cooperate in the cell killing by chemotherapeutic drugs. A few BH3 mimetics are currently being tested in clinical trials of hematologic and solid tumors. Nevertheless, the knowledge about the cellular and molecular mechanisms that regulate responsiveness to BH3 therapy has to be further expanded and will benefit from recent advances in cytometric quantitative technologies.
Topics: Apoptosis; BH3 Interacting Domain Death Agonist Protein; Mitochondria; Models, Biological; Signal Transduction; bcl-2-Associated X Protein
PubMed: 19899133
DOI: 10.1002/cyto.a.20819 -
F1000Research 2020Bax and Bak, two functionally similar, pro-apoptotic proteins of the Bcl-2 family, are known as the gateway to apoptosis because of their requisite roles as effectors of... (Review)
Review
Bax and Bak, two functionally similar, pro-apoptotic proteins of the Bcl-2 family, are known as the gateway to apoptosis because of their requisite roles as effectors of mitochondrial outer membrane permeabilization (MOMP), a major step during mitochondria-dependent apoptosis. The mechanism of how cells turn Bax/Bak from inert molecules into fully active and lethal effectors had long been the focal point of a major debate centered around two competing, but not mutually exclusive, models: direct activation and indirect activation. After intensive research efforts for over two decades, it is now widely accepted that to initiate apoptosis, some of the BH3-only proteins, a subclass of the Bcl-2 family, directly engage Bax/Bak to trigger their conformational transformation and activation. However, a series of recent discoveries, using previously unavailable CRISPR-engineered cell systems, challenge the basic premise that undergirds the consensus and provide evidence for a novel and surprisingly simple model of Bax/Bak activation: the membrane (lipids)-mediated spontaneous model. This review will discuss the evidence, rationale, significance, and implications of this new model.
Topics: Apoptosis; Humans; Proto-Oncogene Proteins c-bcl-2; bcl-2 Homologous Antagonist-Killer Protein; bcl-2-Associated X Protein
PubMed: 32802314
DOI: 10.12688/f1000research.25607.1